The advent of recombinant DNA technology together with DNA mediated gene transfer methods permits introduction of virtually any DNA sequence into mammalian cells. Since there are a large number of human genetic diseases which are due to single gene disorders, the gene transfer systems raised the possibility of gene replacement therapy. In order for gene replacement therapy to be a reality, it is necessary to have highly efficient methods of introducing genes into primary cells. In addition it is now clear that proper regulation of several, if not all, exogenously introduced mammalian genes is dependent upon their chromosomal location. As such, methods to detect and enhance homologous recombination need to be developed before gene replacement therapy can be attempted. We propose a series of experiments which are aimed at these questions. Plasmid DNA containing selectable genes such as bacterial neomycin resistance gene or herpes viral thymidine kinase gene and non-selectable genes such as human globin genes will be used in these experiments. Homologous recombination and gene conversion will be examined by introducing deletion plasmids into mammalian cells and homologous recombination events will be selected. Parameters that affect the rate of homologous recombination will be examined. Based upon certain observations in yeast, specific methods, such as DNA double strand breaks will be tested for their enhancing affect on recombination. Legitimate recombination between chromosomal genes and their exogenously introduced counterparts will also be examined. Finally experiments to develop an in vitro system for recombination are proposed.